The neural crest is a multipotent population of embryonic cells with the ability to contribute to a variety of adult derivatives: neural, endocrine, pigment, and mesenchymal. Because neural crest cells have this intrinsic multipotency, they represent a system in which to explore molecular requirements of multipotent stem cells. Stem cells are a unique tissue with the potential for disease therapy and cell transplantation. However, in order to take advantage of these biological possibilities, we must understand the genetic pathways involved, identify stem cell progenitors prospectively and control their maintenance and differentiation. We have identified a transcription factor, Foxd3 that is sufficient to specify neural crest. Foxd3 is required for normal murine development and the establishment of two embryo-derived stem cell lineages: embryonic stem (ES) cells and trophoblast stem (TS) cells. Aim 1 tests the requirement for Foxd3 in vivo to specify and/or maintain neural crest cells. Adult progenitor cells reside in specialized niches and are recruited to renew or repair tissue thereby maintaining homeostasis. Multipotent neural crest stem cells (NCSCs) can be isolated from both embryonic neural crest cells and some postnatal neural crest derivatives. Foxd3 is expressed in NCSCs and in scattered cells of the adult nervous system known to contain progenitor cells. Experiments in Aim 2 will determine the identity of Foxd3 expressing cells in the adult and if these cells are residing in a stem cell niche. Because Foxd3 is required in other stem cell populations, experiments in Aim 3 will determine if Foxd3 plays a role in the establishment and maintenance of postnatally derived neural crest stem cells (NCSCs). These experiments are all linked to the same biological process of establishing and/or maintaining multipotent cell properties in stem cells from different sources. Understanding the molecular regulation of common properties of different stem cell types is critical to understanding how these cells can maintain multipotency as well as how their differentiation might be controlled in vitro and potentially in vivo.